Compounds and compositions for treating chemical warfare agent-induced injuries

ABSTRACT

Compounds and compositions for treating injuries caused by exposure to chemical warfare agents are described herein.

CLAIM OF PRIORITY

This application is a continuation of U.S. application Ser. No.13/661,812, filed Oct. 26, 2012, which is a continuation of U.S.application Ser. No. 12/466,195, filed May 14, 2009, which claimspriority to U.S. Ser. No. 61/127,722, filed May 14, 2008, and U.S. Ser.No. 61/082,809, filed Jul. 22, 2008, each of which is incorporatedherein by reference in its entirety.

BACKGROUND

The invention relates to compounds and compositions useful for treatinginjuries caused by chemical warfare and similar agents.

A variety of ion channel proteins exist to mediate ion flux acrosscellular membranes. The proper expression and function of ion channelproteins is essential for the maintenance of cell function andintracellular communication. Numerous diseases and disorders are theresult of misregulation of membrane potential or aberrant calciumhandling. Given the central importance of ion channels in modulatingmembrane potential and ion flux in cells, identification of agents thatcan promote or inhibit particular ion channels are of great interest,both as research tools and as therapeutic agents.

SUMMARY OF THE INVENTION

The present invention provides compounds and compositions for treatingor preventing injuries resulting from chemical warfare agents bymodulating the activity of the TRPA1 channel.

In one aspect, the invention features methods for treating injuriescaused by exposure to a chemical warfare agent, or a similar agent, byadministering a compound of formula (I) or a salt thereof,

Wherein each of R¹ and R² is, independently, H, C₁-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, each of which is optionally substituted with1-4 R⁵;L is NR⁶SO₂, SO₂NR⁶, C(O)NR⁶, NR⁶C(O), OC(O)NR⁶, NR⁶C(O)O, NR⁶C(O)NR⁶,S, S(O), S(O)₂, NR⁶, CH₂, O, C(O)NS(O)₂, S(O)₂NC(O), heteroaryl, orcyclyl;R³ is C₄-C₁₄ cyclyl, heterocyclyl, aryl, or heteroaryl, each of which isoptionally substituted with 1-4 R⁷;each R⁵ is independently halo, hydroxyl, alkoxy, thiol, alkylthio,amino, alkylamino, dialkylamino, cyano, nitro, amido, alkylamido,dialkylamido, thioyl, sulfonyl, cyclyl, heterocyclyl, aryl, orheteroaryl;each R⁶ is independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl,aryl, arylalkyl, or acetyl;each R⁷ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl,cyclyl, heterocyclyl, aryl, heteroaryl, halo, hydroxyl, alkoxy, thiol,alkylthio, aryloxy, arylalkoxy, amino, akylamino, dialkylamino, thioyl,alkylthioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylurea, hydroxylalkoxyl, alkoxy alkoxyl, acyl, nitro, or cyano, each of which isoptionally substituted with 1-3 R⁸;each R⁸ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, thiol, alkylthio, aryloxy, amino, akylamino,dialkylamino, thioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylureaacyl, nitro, cyano, cyclyl, heterocyclyl, aryl, or heteroaryl;R⁹ is independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, aryloxy, arylalkoxy, amino, akylamino,dialkylamino, thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido, urea,sulfonylurea, acyl, nitro, cyano, and is optionally substituted with 1-3R⁸;each of R¹¹-R¹⁴ is, independently, H, C₁-C₆ alkyl, C₂-C₆ alkenyl, orC₂-C₆ alkynyl, halo, hydroxyl, alkoxy, amino, akylamino, thiol,alkylthiol, nitro, or cyano, each of which is optionally substitutedwith 1-2 R⁸;each of m and n is, independently, 0, 1, 2, 3, 4, 5, or 6.

In some embodiments, R³ is aryl, such as phenyl. In some embodiments, R³is a nitrogen containing heterocyclyl such as

In other embodiments, R³ is a nitrogen containing heteroaryl, forexample,

In some embodiments, the heteroaryl is substituted by R⁷ is in the 3 or4 position of the 5 membered ring. In some embodiments, R³ is a 6membered nitrogen-containing heteroaryl, for example,

In some embodiments, R³ is a heteroaryl or heterocycyl having two fusedrings. In some embodiments, R³ is a heteroaryl or heterocycyl havingthree fused rings.

In some embodiments, R³ is substituted by 1-3 R⁷. In some embodiments,R⁷ is Me, OMe, or halo.

In another aspect, the invention relates to a method for treatinginjuries caused by exposure to chemical warfare agents by administeringan effective amount of a compound of Formula II, or a salt thereof:

wherein R3 is as defined above and n is 1-3.

In another aspect, the invention relates to a method for treatinginjuries caused by exposure to chemical warfare agents by administeringan effective amount of a compound of Formula III, or a salt thereof:

Wherein each of R²¹ and R²² is, independently, H, C₁-C₆ alkyl, C₂-C₆alkenyl, or C₂-C₆ alkynyl, each of which is optionally substituted with1-4 R²⁵;L is NR²⁶SO₂, SO₂NR²⁶, C(O)NR²⁶, NR²⁶C(O), OC(O)NR²⁶, NR²⁶C(O)O,NR²⁶C(O)NR²⁶, S, S(O), S(O)₂, NR²⁶, CH₂, O, C(O)NS(O)₂, S(O)₂NC(O),heteroaryl, or cyclyl;R²³ is C₄-C₁₄ cyclyl, heterocyclyl, aryl, or heteroaryl, each of whichis optionally substituted with 1-4 R²⁷;each R²⁵ is independently halo, hydroxyl, alkoxy, thiol, alkylthio,amino, alkylamino, dialkylamino, cyano, nitro, amido, alkylamido,dialkylamido, thioyl, sulfonyl, cyclyl, heterocyclyl, aryl, orheteroaryl;each R²⁶ is independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl,aryl, arylalkyl, or acetyl;each R²⁷ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl,cyclyl, heterocyclyl, aryl, heteroaryl, halo, hydroxyl, alkoxy, thiol,alkylthio, aryloxy, arylalkoxy, amino, akylamino, dialkylamino, thioyl,alkylthioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylurea, hydroxylalkoxyl, alkoxy alkoxyl, acyl, nitro, or cyano, each of which isoptionally substituted with 1-3 R²⁸;each R²⁸ is independently C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, thiol, alkylthio, aryloxy, amino, akylamino,dialkylamino, thioyl, sulfonyl, sulfonamidyl, amido, urea, sulfonylureaacyl, nitro, cyano, cyclyl, heterocyclyl, aryl, or heteroaryl;R²⁹ is independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, aryloxy, arylalkoxy, amino, akylamino,dialkylamino, thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido, urea,sulfonylurea, acyl, nitro, cyano, and is optionally substituted with 1-3R²⁸;each of R³¹-R³⁴ is, independently, H, C₁-C₆ alkyl, C₂-C₆ alkenyl, orC₂-C₆ alkynyl, halo, hydroxyl, alkoxy, amino, akylamino, thiol,alkylthiol, nitro, or cyano, each of which is optionally substitutedwith 1-2 R²⁸;each of m and n is, independently, 0, 1, 2, 3, 4, 5, or 6.

In another aspect, the present invention provides a method for treatinginjuries from chemical warfare agents by administering an effectiveamount of a compound of Formula IV, or a salt thereof:

Wherein each of R⁴¹ and R⁴² is, independently, H or C₁-C₆ alkyl; one ofR⁴³ and R⁴⁴ is H and the other is M_(m)R⁵³; R⁵³ is substituted orunsubstituted aryl; M, independently for each occurrence, isC(R⁶¹)(R⁶²), NR₁, O, S, C(O), S(O), or S(O₂), preferably selected suchthat no two heteroatoms are adjacent to each other, where each of R⁶¹and R⁶² is, independently, H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆alkynyl, halo, hydroxyl, alkoxy, amino, akylamino, thiol, alkylthiol,nitro, or cyano; R⁴⁵ is H, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl,halo, hydroxyl, alkoxy, aryloxy, arylalkoxy, amino, akylamino,dialkylamino, thioyl, alkylthioyl, sulfonyl, sulfonamidyl, amido, urea,sulfonylurea, acyl, nitro, or cyano; andm is an integer from 1-5.

In some embodiments, R⁴¹ and R⁴² are both methyl. In some embodiments,R⁴⁵ is H. In some embodiments, M is CH₂—C(O)NH—.

In one aspect, the invention features a method of treating a subject whohas been exposed to a chemical warfare agent, the method comprisingadministering to a subject an effective amount of a compound of Formula(I), (II), (III), or (IV), or a salt thereof. In some embodiments thetreatment reduces the severity of injury resulting from the exposure tothe chemical warfare agent.

In some embodiments, the compound is administered orally, viaintramuscular injection, by topical ocular administration, topically, orby inhalation. In some embodiments, the subject being treated is ahuman.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1 and 2 are Tables showing exemplary compounds.

DETAILED DESCRIPTION OF THE INVENTION Definitions

The term “acyl” refers to a group represented by the general formulahydrocarbylC(O)—, preferably alkylC(O)—.

The term “acylamino” refers to a moiety that can be represented by thegeneral formula:

wherein R₉ is as defined below, and R′11 represents a hydrogen, analkyl, an alkenyl or —(CH2)m-R8, where m and R8 are as defined herein.

The term “aliphatic group” refers to a straight-chain, branched-chain,or cyclic aliphatic hydrocarbon group and includes saturated andunsaturated aliphatic groups, such as an alkyl group, an alkenyl group,and an alkynyl group.

The term “alkenyl,” as used herein, refers to an aliphatic groupcontaining at least one double bond.

The terms “alkoxyl” or “alkoxy” as used herein refers to an alkyl group,as defined below, having an oxygen radical attached thereto.Representative alkoxyl groups include methoxy, ethoxy, propyloxy,tert-butoxy and the like. An “ether” is two hydrocarbons covalentlylinked by an oxygen.

The term “alkyl” refers to the radical of saturated aliphatic groups,including straight-chain alkyl groups, branched-chain alkyl groups,cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, andcycloalkyl-substituted alkyl groups. In preferred embodiments, astraight chain or branched chain alkyl has 30 or fewer carbon atoms inits backbone (e.g., C1-C30 for straight chains, C3-C30 for branchedchains), and more preferably 20 or fewer, and most preferably 10 orfewer. Likewise, preferred cycloalkyls have from 3-10 carbon atoms intheir ring structure, and more preferably have 5, 6 or 7 carbons in thering structure.

The term “alkynyl”, as used herein, refers to an aliphatic groupcontaining at least one triple bond.

The term “alkylthio” refers to a hydrocarbyl group having a sulfurradical attached thereto. In some embodiments, the “alkylthio” moiety isrepresented by one of —S-alkyl, —S-alkenyl, or —S-alkynyl.Representative alkylthio groups include methylthio, ethylthio, and thelike.

The terms “amine” and “amino” refer to both unsubstituted andsubstituted amines, e.g., a moiety that can be represented by thegeneral formula:

wherein R₉, R₁₀ and R′₁₀ each independently represent a hydrogen, analkyl, an alkenyl, —(CH₂)_(m)—R₈, or R₉ and R₁₀ taken together with theN atom to which they are attached complete a heterocycle having from 4to 8 atoms in the ring structure; R₈ represents an aryl, a cycloalkyl, acycloalkenyl, an alkoxy, a heterocycle or a polycycle; and m is zero oran integer in the range of 1 to 8.

The term “amido” refers to a moiety that can be represented by thegeneral formula:

wherein R₉, R₁₀ are as defined above.

The term “aralkyl”, as used herein, refers to an alkyl group substitutedwith an aryl group (e.g., an aromatic or heteroaromatic group).

The term “aryl” as used herein includes 5-, 6-, and 7-memberedsingle-ring aromatic groups that may include from zero to fourheteroatoms, for example, benzene, pyrrole, furan, thiophene, imidazole,oxazole, thiazole, triazole, pyrazole, pyridine, pyrazine, pyridazineand pyrimidine, and the like. Those aryl groups having heteroatoms inthe ring structure may also be referred to as “aryl heterocycles” or“heteroaromatics.” The aromatic ring can be substituted at one or morering positions with such substituents as described above, for example,halogen, azide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl,polycyclyl, hydroxyl, alkoxyl, amino, nitro, sulfhydryl, imino, amido,phosphate, phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether,alkylthio, sulfonyl, sulfonamido, ketone, aldehyde, ester, heterocyclyl,aromatic or heteroaromatic moieties, —CF3, —CN, or the like. The term“aryl” also includes polycyclic ring systems having two or more cyclicrings in which two or more carbons are common to two adjoining rings(the rings are “fused rings”) wherein at least one of the rings isaromatic, e.g., the other cyclic rings can be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls and/or heterocyclyls.

The term “carbocycle or cyclyl,” as used herein, refers to an aromaticor non-aromatic ring in which each atom of the ring is carbon.

The term “carbonyl” refers to moieties represented by the generalformula:

wherein X is a bond or represents an oxygen or a sulfur, and R₁₁represents a hydrogen, an alkyl, an alkenyl, —(CH₂)_(m)—R₈ or apharmaceutically acceptable counter-ion, R′₁₁ represents a hydrogen, analkyl, an alkenyl or —(CH₂)_(m)—R₈, where m and R₈ are as defined above.Where X is an oxygen and R₁₁ or R′₁₁ is not hydrogen, the formularepresents an “ester”. Where X is an oxygen, and R₁₁ is as definedabove, the moiety is referred to herein as a carboxyl group, andparticularly when R₁₁ is a hydrogen, the formula represents a“carboxylic acid”. Where X is an oxygen, and R′₁₁ is hydrogen, theformula represents a “formate”. In general, where the oxygen atom of theabove formula is replaced by sulfur, the formula represents a“thiocarbonyl” group. Where X is a sulfur and R₁₁ or R′₁₁ is nothydrogen, the formula represents a “thioester.” Where X is a sulfur andR₁₁ is hydrogen, the formula represents a “thiocarboxylic acid.” Where Xis a sulfur and R₁₁′ is hydrogen, the formula represents a“thioformate.” On the other hand, where X is a bond, and R₁₁ is nothydrogen, the above formula represents a “ketone” group. Where X is abond, and R₁₁ is hydrogen, the above formula represents an “aldehyde”group.

The term “ester”, as used herein, refers to a group —C(O)OR⁹ wherein R⁹represents a hydrocarbyl group.

The terms “halo” and “halogen” as used herein means halogen and includeschloro, fluoro, bromo, and iodo.

The terms “hetaralkyl” and “heteroaralkyl”, as used herein, refers to analkyl group substituted with a hetaryl group.

The terms “heterocyclyl” or “heterocyclic group” refer to 3- to10-membered ring structures, more preferably 3- to 7-membered rings,whose ring structures include one to four heteroatoms. Heterocycles canalso be polycycles. The term “heterocyclyl” or “heterocyclic group”includes “heteroaryl” and “saturated or partially saturatedheterocyclyl” structures. The term “heteroaryl” refers to an aromatic5-8 membered monocyclic, 8-12 membered bicyclic, or 11-14 memberedtricyclic ring system having 1-3 heteroatoms if monocyclic, 1-6heteroatoms if bicyclic, or 1-9 heteroatoms if tricyclic, saidheteroatoms selected from O, N, or S (e.g., carbon atoms and 1-3, 1-6,or 1-9 heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic,respectively). Any ring atom can be substituted (e.g., by one or moresubstituents). The term “saturated or partially saturated heterocyclyl”refers to abnon-aromatic cylic structure that includes at least oneheteroatom. Heterocyclyl groups include, for example, thiophene,thianthrene, furan, pyran, isobenzofuran, chromene, xanthene,phenoxathiin, pyrrole, imidazole, pyrazole, isothiazole, isoxazole,pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole,indole, indazole, purine, quinolizine, isoquinoline, quinoline,phthalazine, naphthyridine, quinoxaline, quinazoline, cinnoline,pteridine, carbazole, carboline, phenanthridine, acridine, pyrimidine,phenanthroline, phenazine, phenarsazine, phenothiazine, furazan,phenoxazine, pyrrolidine, oxolane, thiolane, oxazole, piperidine,piperazine, morpholine, lactones, lactams such as azetidinones andpyrrolidinones, sultams, sultones, and the like. The heterocyclic ringcan be substituted at one or more positions with such substituents asdescribed above, as for example, halogen, alkyl, aralkyl, alkenyl,alkynyl, cycloalkyl, hydroxyl, amino, nitro, sulfhydryl, imino, amido,phosphate, phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether,alkylthio, sulfonyl, ketone, aldehyde, ester, a heterocyclyl, anaromatic or heteroaromatic moiety, —CF₃, —CN, or the like.

The term “heterocyclylalkyl”, as used herein, refers to an alkyl groupsubstituted with a heterocycle group.

The term “hydrocarbyl”, as used herein, refers to a group that is bondedthrough a carbon atom that does not have a ═O or ═S substituent, andtypically has at least one carbon-hydrogen bond and a primarily carbonbackbone, but may optionally include heteroatoms. Thus, groups likemethyl, ethoxyethyl, 2-pyridyl, and trifluoromethyl are considered to behydrocarbyl for the purposes of this application, but substituents suchas acetyl (which has a ═O substituent on the linking carbon) and ethoxy(which is linked through oxygen, not carbon) are not. Hydrocarbyl groupsinclude, but are not limited to aryl, heteroaryl, carbocycle,heterocycle, alkyl, alkenyl, alkynyl, and combinations thereof.

As used herein, the term “nitro” means —NO2; the term “halogen”designates —F, —Cl, —Br or —I; the term “sulfhydryl” means —SH; the term“hydroxyl” means —OH; and the term “sulfonyl” means —SO2-.

The terms “polycyclyl” or “polycyclic group” refer to two or more rings(e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls and/orheterocyclyls) in which two or more carbons are common to two adjoiningrings, e.g., the rings are “fused rings”. Rings that are joined throughnon-adjacent atoms are termed “bridged” rings. Each of the rings of thepolycycle can be substituted with such substituents as described above,as for example, halogen, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl,hydroxyl, amino, nitro, sulfhydryl, imino, amido, phosphate,phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether, alkylthio,sulfonyl, ketone, aldehyde, ester, a heterocyclyl, an aromatic orheteroaromatic moiety, —CF3, —CN, or the like.

Exemplary monocyclic rings include furan, thiophene, pyrrole, pyrroline,pyrrolodine, oxazole, thiazole, imidazole, imidazoline, pyrazole,pyrazoline, pyrazolidine, isoxazole, isothiazole, oxadiazole, triazole,thiadiazole, pyran, pyridine, piperidine, dioxane, morpholine, dithiane,thiomorpholine, pyridazine, pyrimidine, pyrazine, piperazine, triazine,and trithiane.

Exemplary bicyclic rings include indolizinyl, indolyl, isoindolyl,indolinyl, benzofuranyl, benzothiophenyl, indazolyl, benzimidazolyl,benthiazolyl, purinyl, quinolizinyl, quinolinyl, isoquinolinyl,cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, naphthyridinyl,pteridinyl, indenyl, naphthalenyl, azulenyl, imidazopyridazionyl,pyrazolopyrimidinedionyl, or pyrrolopyrimidinedionyl moieties.

Exemplary tricyclic rings include carbazole, acridine, phenazine,phenothiazine, phenoxazine, fluorine, and anthracene.

The phrase “protecting group” as used herein means temporarysubstituents which protect a potentially reactive functional group fromundesired chemical transformations. Examples of such protecting groupsinclude esters of carboxylic acids, silyl ethers of alcohols, andacetals and ketals of aldehydes and ketones, respectively. The field ofprotecting group chemistry has been reviewed (Greene, T. W.; Wuts, P. G.M. Protective Groups in Organic Synthesis, 2^(nd) ed.; Wiley: New York,1991).

The term “substituted” refers to moieties having substituents replacinga hydrogen on one or more carbons of the backbone. It will be understoodthat “substitution” or “substituted with” includes the implicit provisothat such substitution is in accordance with permitted valence of thesubstituted atom and the substituent, and that the substitution resultsin a stable compound, e.g., which does not spontaneously undergotransformation such as by rearrangement, cyclization, elimination, etc.As used herein, the term “substituted” is contemplated to include allpermissible substituents of organic compounds. In a broad aspect, thepermissible substituents include acyclic and cyclic, branched andunbranched, carbocyclic and heterocyclic, aromatic and non-aromaticsubstituents of organic compounds. The permissible substituents can beone or more and the same or different for appropriate organic compounds.For purposes of this invention, the heteroatoms such as nitrogen mayhave hydrogen substituents and/or any permissible substituents oforganic compounds described herein which satisfy the valences of theheteroatoms. Substituents can include any substituents described herein,for example, a halogen, a hydroxyl, a carbonyl (such as a carboxyl, analkoxycarbonyl, a formyl, or an acyl), a thiocarbonyl (such as athioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, aphosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine,an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, asulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, aheterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. Itwill be understood by those skilled in the art that the moietiessubstituted on the hydrocarbon chain can themselves be substituted, ifappropriate. For instance, the substituents of a substituted alkyl mayinclude substituted and unsubstituted forms of amino, azido, imino,amido, phosphoryl (including phosphonate and phosphinate), sulfonyl(including sulfate, sulfonamido, sulfamoyl and sulfonate), and silylgroups, as well as ethers, alkylthios, carbonyls (including ketones,aldehydes, carboxylates, and esters), —CF₃, —CN and the like. Exemplarysubstituted alkyls are described below. Cycloalkyls can be furthersubstituted with alkyls, alkenyls, alkoxys, alkylthios, aminoalkyls,carbonyl-substituted alkyls, —CF₃, —CN, and the like. Analogoussubstitutions can be made to alkenyl and alkynyl groups to produce, forexample, aminoalkenyls, aminoalkynyls, amidoalkenyls, amidoalkynyls,iminoalkenyls, iminoalkynyls, thioalkenyls, thioalkynyls,carbonyl-substituted alkenyls or alkynyls.

The term “sulfate” refers to a moiety that can be represented by thegeneral formula:

in which R41 is as defined herein.

The term “sulfonamido” refers to a moiety that can be represented by thegeneral formula:

in which R9 and R′11 are as defined above.

The term “sulfonate” refers to a moiety that can be represented by thegeneral formula:

in which R41 is an electron pair, hydrogen, alkyl, cycloalkyl, or aryl.

The terms “sulfoxido” or “sulfinyl,” as used herein, refers to a moietythat can be represented by the general formula —S(═O)—R44, in which R44is selected from the group consisting of hydrogen, alkyl, alkenyl,alkynyl, cycloalkyl, heterocyclyl, aralkyl, or aryl.

The term “thioester,” as used herein, refers to a group —C(O)SR⁹ or—SC(O)R⁹ wherein R⁹ represents a hydrocarbyl.

As used herein, the definition of each expression, e.g., alkyl, m, n,etc., when it occurs more than once in any structure, is intended to beindependent of its definition elsewhere in the same structure.

The terms “antagonist” and “inhibitor” are used interchangeably to referto an agent that decreases or suppresses a biological activity, such asto repress an activity of an ion channel, such as TRPA1. TRPA1inhibitors include inhibitors having any combination of the structuraland/or functional properties disclosed herein.

An “effective amount” of, e.g., a TRPA1 antagonist, with respect to thesubject methods of treatment, refers to an amount of the antagonist in apreparation which, when applied as part of a desired dosage regimenbrings about a desired clinical or functional result. Without beingbound by theory, an effective amount of a TRPA1 antagonist for use inthe methods of the present invention, includes an amount of a TRPA1antagonist effective to decrease one or more in vitro or in vivofunctions of a TRPA1 channel. Exemplary functions include, but are notlimited to, membrane polarization (e.g., an antagonist may promotehyperpolarization of a cell), ion flux, ion concentration in a cell,outward current, and inward current. Compounds that antagonize TRPA1function include compounds that antagonize an in vitro or in vivofunctional activity of TRPA1. When a particular functional activity isonly readily observable in an in vitro assay, the ability of a compoundto inhibit TRPA1 function in that in vitro assay serves as a reasonableproxy for the activity of that compound. In certain embodiments, aneffective amount is an amount sufficient to inhibit a TRPA1-mediatedcurrent and/or the amount sufficient to inhibit TRPA1 mediated ion flux.

The term “preventing,” when used in relation to a condition, such as alocal recurrence (e.g., pain), a disease such as cancer, a syndromecomplex such as heart failure or any other medical condition, is wellunderstood in the art, and includes administration of a compositionwhich reduces the frequency of, or delays the onset of, symptoms of amedical condition or injury in a subject relative to a subject whichdoes not receive the composition. Thus, prevention of cancer includes,for example, reducing the number of detectable cancerous growths in apopulation of patients receiving a prophylactic treatment relative to anuntreated control population, and/or delaying the appearance ofdetectable cancerous growths in a treated population versus an untreatedcontrol population, e.g., by a statistically and/or clinicallysignificant amount. Prevention of an infection includes, for example,reducing the number of diagnoses of the infection in a treatedpopulation versus an untreated control population, and/or delaying theonset of symptoms of the infection in a treated population versus anuntreated control population. Prevention of pain includes, for example,reducing the magnitude of, or alternatively delaying, pain sensationsexperienced by subjects in a treated population versus an untreatedcontrol population.

The term “prodrug” is intended to encompass compounds that, underphysiological conditions, are converted into the therapeutically activeagents of the present invention. A common method for making a prodrug isto include selected moieties that are hydrolyzed under physiologicalconditions to reveal the desired molecule. In other embodiments, theprodrug is converted by an enzymatic activity in the host animal.

Exemplary compounds are shown in Table 1 below:

“A” represents a compound demonstrating activity of <1 μM as measured inthe patch clamp assay described in Example 3. “B” represents a compounddemonstrating activity of >1 μM-<10 μM as measured in the patch clampassay. “C” represents a compound demonstrating activity of >10 μM asmeasured in the patch clamp assay. “D” represents other exemplarycompounds.

TABLE 1 1 2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3-D methoxyphenyl)acetamide 22-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Cfluorophenyl)acetamide 32-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- Cp-tolylacetamide 4(E)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N′-(4- B(trifluoromethyl)benzylidene)acetohydrazide 52-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-methyl- DN-(2-(pyridin-2-yl)ethyl)acetamide 62-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- D(pyridine-2-yl)ethyl)acetamide 72-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3- Dmethoxyphenyl)acetamide 82-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Aadamantylethyl)acetamide 92-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5- Bfluoro-2-methylphenyl)acetamide 10N-cyclooctyl-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamide 112-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(adamantylmethyl)acetamide 122-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(thiochroman-4-yl)acetamide 132-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Dfluorophenyl)acetamide 142-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- D(furan-2-yl)-2-(pyrrolidin-1-yl)ethyl)acetamide 152-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(3-fluoro-4-methoxybenzyl)-N-methylacetamide 162-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(3-methoxybenzyl)acetamide 172-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(4-fluorophenethyl)acetamide 182-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-p- Atolylpropanamide 19N-((2,3-dihydrobenzo[b][1,4]dioxin-2-yl)methyl)-2-(1,3-dimethyl-2,6- Ddioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide 202-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- A(2,3-dihydro-1H-inden-5-yl)acetamide 212-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(2,3-dimethylcyclohexyl)acetamide 22 methyl4-((2-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin- D7(6H)-yl)acetamido)methyl)benzoate 23N-(3,4-dimethoxybenzyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 242-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- Disobutyl-N-(dioxytetrahydrothiophen-3-yl)acetamide 25N-((3,5-dimethyl-1-phenyl-1H-pyrazol-4-yl)methyl)-2-(1,3-dimethyl- D2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-methylacetamide 262-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- Dmethyl-N-(4-methylbenzyl)acetamide 272-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- Dcyclohexyl-N-ethylacetamide 282-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(3,4-dihydro-2H-benzo[b][1,4]dioxepin-7-yl)acetamide 292-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(1-cyclopentyl-1H-pyrazol-5-yl)acetamide 302-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- Dmethyl-N-(1,2,3,4-tetrahydronaphthalen-1-yl)acetamide 312-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(4-phenylbutan-2-yl)acetamide 32N-cyclohexyl-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)-N-ethylacetamide 332-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- A(2-(4-chlorophenylthio)ethyl)acetamide 34N-((2,3-dihydrobenzo[b][1,4]dioxin-2-yl)methyl)-2-(1,3-dimethyl-2,6- Ddioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-methylacetamide 35N-(2-cyclohexenylethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 362-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(1-(5,6,7,8-tetrahydronaphthalen-2-yl)ethyl)acetamide 37N-(3,5-dichloropyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- D1H-purin-7(6H)-yl)propanamide 38N-(5-chloropyridin-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)propanamide 39N-((4-chlorophenyl)(cyclopropyl)methyl)-2-(1,3-dimethyl-2,6-dioxo- D2,3-dihydro-1H-purin-7(6H)-yl)acetamide 40N-(1-(3,4-dihydro-2H-benzo[b][1,4]dioxepin-7-yl)-2-methylpropyl)-2- D(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide 41N-((1-benzyl-1H-pyrazol-4-yl)methyl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Ddihydro-1H-purin-7(6H)-yl)-N-methylacetamide 422-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2,2- Cdiphenylpropyl)acetamide 432-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-((2- Dethylbenzofuran-3-yl)methyl)-N-methylacetamide 44N-(cyclohexylmethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 45N-cyclohexyl-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)-N-methylpropanamide 463-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Cmethoxybenzyl)propanamide 472-(1,3-diethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Cmethoxyphenethyl)acetamide 481-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Cmethoxyphenethyl)methanesulfonamide 49N-(2-cyclohexylethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Cpurin-7(6H)-yl)acetamide 502-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(4- Chydroxycyclohexyl)ethyl)acetamide 512-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3,4- Bdimethylphenethyl)acetamide 52N-(2-(biphenyl-4-yl)ethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Apurin-7(6H)-yl)acetamide 53N-(4-(benzyloxy)phenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- B1H-purin-7(6H)-yl)acetamide 542-(1,3-dimethyl-2,6,8-trioxo-2,3,7,8-tetrahydro-1H-purin-9(6H)-yl)-N- C(4-methoxyphenethyl)acetamide 552-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- C((3R,4R)-quinuclidin-3-yl)acetamide 56N-(4-butoxyphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 57N-(4-cyclohexylphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Apurin-7(6H)-yl)acetamide 582-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-((1S,2R)- B2-(4-methoxyphenyl)cyclopropyl)acetamide 59N-(4-tert-butylphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 602-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Bmethoxybenzylsulfonyl)acetamide 61N-(4-methoxyphenethyl)-2-(1,3,9-trimethyl-2,6,8-trioxo-2,3-dihydro- B1H-purin-7(6H,8H,9H)-yl)acetamide 62 4-methylphenethyl2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin- B 7(6H)-yl)acetate 632-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- C(piperidin-1-yl)ethyl)acetamide 642-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Cmorpholinoethyl)acetamide 652-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-((1S,2S)- B2-(4-methoxyphenyl)cyclopropyl)acetamide 662-(1,3-dicyclopropyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Dmethoxyphenethyl)acetamide 672-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(4- Dmethoxycyclohexyl)ethyl)acetamide 682-(1,3-dimethyl-2,6,8-trioxo-2,3-dihydro-1H-purin-7(6H,8H,9H)-yl)-N- C(4-methoxyphenethyl)acetamide 692-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- C(pyridine-4-yl)ethyl)acetamide 702-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(4- Bmethylpiperazin-1-yl)ethyl)acetamide 711,3-dimethyl-7-(2-(4-methylphenethylamino)ethyl)-1H-purine- B2,6(3H,7H)-dione 72N-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)ethyl)-N- B(4-methylphenethyl)methanesulfonamide 732-(3-methyl-2,6-dioxo-1-propyl-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Cmethylphenethyl)acetamide 741,3-dimethyl-7-(2-(methyl(4-methylphenethyl)amino)ethyl)-1H-purine- B2,6(3H,7H)-dione 752-(1-methyl-2,6-dioxo-3-propyl-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Bmethylphenethyl)acetamide 76N-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)ethyl)-N- A(4-methylphenethyl)acetamide 772-(1-methyl-2,6-dioxo-3-propyl-2,3-dihydro-1H-purin-7(6H)-yl)-N- Aphenethylacetamide 782-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Bphenylpropyl)acetamide 792-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(3- Bfluorophenethyl)acetamide 802-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Bethylphenethyl)acetamide 81(S)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(1- Bhydroxy-3-phenylpropan-2-yl)acetamide 82N-(2,3-dimethoxyphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- B1H-purin-7(6H)-yl)acetamide 83N-(2,3-dichlorophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 84N-(2-(benzo[d][1,3]dioxol-5-yl)ethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Bdihydro-1H-purin-7(6H)-yl)acetamide 85(R)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(1- Bhydroxy-3-phenylpropan-2-yl)acetamide 86N-(2,5-dimethoxyphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- B1H-purin-7(6H)-yl)acetamide 87N-(2,4-dichlorophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 88N-(2,6-dichlorophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 89N-(2-bromophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Bpurin-7(6H)-yl)acetamide 902-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- D(quinuclidin-3-yl)ethyl)acetamide 912-(3-(2-(dimethylamino)ethyl)-1-methyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)-N-(4-methylphenethyl)acetamide 922-(1-(2-(dimethylamino)ethyl)-3-methyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)-N-(4-methylphenethyl)acetamide 93N-(6-chlorobenzo[d]thiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Ddihydro-1H-purin-7(6H)-yl)acetamide 94N-(6-bromobenzo[d]thiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Adihydro-1H-purin-7(6H)-yl)acetamide 95N-(4-chlorobenzo[d]thiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Ddihydro-1H-purin-7(6H)-yl)acetamide 962-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(5,6- Ddimethylbenzo[d]thiazol-2-yl)acetamide 972-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6- Aethoxybenzo[d]thiazol-2-yl)acetamide 982-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Dmethoxybenzo[d]thiazol-2-yl)acetamide 992-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Dmethylbenzo[d]thiazol-2-yl)acetamide 1002-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(6- D(methylsulfonyl)benzo[d]thiazol-2-yl)acetamide 1012-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Dhydroxy-2-phenylethyl)acetamide 102N-(3-chlorophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 103(S)-2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)-3-phenylpropanamide 104N-(2-chloro-6,7-dimethoxyquinazolin-4-yl)-2-(1,3-dimethyl-2,6-dioxo- D2,3-dihydro-1H-purin-7(6H)-yl)acetamide 105N-(4-bromophenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 1062-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2,2- Ddiphenylethyl)acetamide 107 (S)-methyl2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)-3-phenylpropanoate 108N-(3,5-dimethoxyphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- D1H-purin-7(6H)-yl)acetamide 109(R)-2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)-3-phenylpropanamide 1102-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4,5,6,7- Dtetrahydrobenzo[d]thiazol-2-yl)acetamide 111 ethyl2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)-5-methylthiazole-4-carboxylate 112N-(2-(1,4-diazabicyclo[2.2.2]octan-2-yl)ethyl)-2-(1,3-dimethyl-2,6- Ddioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide 113 ethyl2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)benzo[d]thiazole-6-carboxylate 1142-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-methyl- DN-phenethylacetamide 1151,3-dimethyl-7-((5-(4-methylbenzyl)-1,3,4-oxadiazol-2-yl)methyl)-1H- Dpurine-2,6(3H,7H)-dione 1162-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(6- Dmethylpyridin-3-yl)ethyl)acetamide 1172-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(1- Dmethyl-1H-imidazol-2-yl)ethyl)acetamide 1181,3-dimethyl-7-(3-(6-methyl-1H-indol-2-yl)propyl)-1H-purine- D2,6(3H,7H)-dione 1192-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-ethyl-N- D(pyridine-4-ylmethyl)acetamide 120N-(4,5-dihydrothiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 121 ethyl2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)-4-methylthiazole-5-carboxylate 122N-(4H-chromeno[4,3-d]thiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Ddihydro-1H-purin-7(6H)-yl)acetamide 123 ethyl2-(2-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)- Dyl)acetamido)thiazol-4-yl)acetate 1242-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4- Dmethylthiazol-2-yl)acetamide 1252-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(1,2,3,4- Dtetrahydronaphthalen-1-yl)acetamide 126N-(2,3-dihydro-1H-inden-1-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro- D1H-purin-7(6H)-yl)acetamide 127N′-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetyl)- D4-methylbenzohydrazide 1282-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2-(5- Dmethylpyridin-2-yl)ethyl)acetamide 1291-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)ethyl)-1- D(4-methylphenethyl)urea 1301,3-dimethyl-7-((5-p-tolyl-1,3,4-oxadiazol-2-yl)methyl)-1H-purine- D2,6(3H,7H)-dione 131N-(2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)ethyl)-N- D(4-methylphenethyl)propionamide 1321,3-dimethyl-7-((2-(4-methylphenethyl)cyclopropyl)methyl)-1H- Dpurine-2,6(3H,7H)-dione 133N-(3-bromo-4-methoxyphenethyl)-2-(1,3-dimethyl-2,6-dioxo-2,3- Ddihydro-1H-purin-7(6H)-yl)acetamide 134N-((1R,2R)-1,3-dihydroxy-1-phenylpropan-2-yl)-2-(1,3-dimethyl-2,6- Ddioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetamide 1352-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N- D(piperidin-4-yl)acetamide 1362-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2,2,6,6- Dtetramethylpiperidin-4-yl)acetamide 1372-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(2- Dhydroxy-2-phenylethyl)-N-methylacetamide 1382-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)-N-(4,6- Ddimethylpyrimidin-2-yl)acetamide 139N-benzyl-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)- DN-(4,5,6,7-tetrahydrobenzo[d]thiazol-2-yl)acetamide 140N-benzyl-2-(8-chloro-1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H-purin- D7(6H)-yl)-N-(4,5,6,7-tetrahydrobenzo[d]thiazol-2-yl)acetamide 141N-(4-acetylthiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-2,3-dihydro-1H- Dpurin-7(6H)-yl)acetamide 1422-(8-chloro-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)-N-(4- Dmethylphenethyl)acetamide; 143N-(6-butylbenzo[d]thiazol-2-yl)-2-(8-chloro-1,3-dimethyl-2,6-dioxo- D1,2,3,6-tetrahydropurin-7-yl)acetamide; 1442-(8-chloro-1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)-N-(2- D(adamant-1-yl)ethyl)acetamide; 1451-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-1-(4- Dmethylphenethyl)urea; 146N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-N-(4- Dmethylphenethyl)propionamide; 147N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-N-(4- Dmethylphenethyl)pentanamide; 1482-(3-(2-hydroxyethyl)-1-methyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)-N-(4-methylphenethyl)acetamide; 1492-(3-(2-amino-2-oxoethyl)-1-methyl-2,6-dioxo-1,2,3,6-tetrahydropurin- D7-yl)-N-(4-methylphenethyl)acetamide; 150N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-4- Doxo-4-phenylbutanamide; 151N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-5-(4- Dfluorophenyl)-5-oxopentanamide; 1522-(3-chloro-4-(trifluoromethyl)phenyl)-N-(2-(1,3-dimethyl-2,6-dioxo- D1,2,3,6-tetrahydropurin-7-yl)ethyl)acetamide; 1532-(3-chloro-4-fluorophenyl)-N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6- Dtetrahydropurin-7-yl)ethyl)acetamide; 1541-benzyl-N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)-1H-pyrazole-4-carboxamide; 155N-(2-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethylamino)-2-oxoethyl)-4-methylbenzamide; 156N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-1- Disopropyl-1H-pyrazole-4-carboxamide; 157N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-2- Dphenylacetamide; 158 benzyl2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethylcarbamate; 159N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-4- Dmethylbenzamide; 160N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)ethyl)-3,4,5-D trimethoxybenzamide; 1614-chloro-N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)picolinamide; 1625-bromo-N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)furan-2-carboxamide; 1635-chloro-N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)thiophene-2-carboxamide; 164N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)pyrimidine-4-carboxamide; 165N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)pyrazine-2-carboxamide; 166N-(2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7- Dyl)ethyl)cyclopentanecarboxamide; 167N-(4H-chromeno[4,3-d]oxazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-1,2,3,6- Dtetrahydropurin-7-yl)acetamide 168N-(3,4-dihydrochromeno[3,4-d]imidazol-2-yl)-2-(1,3-dimethyl-2,6- Ddioxo-1,2,3,6-tetrahydropurin-7-yl)acetamide 1692-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)-N-(3-methyl- D3,4-dihydrochromeno[3,4-d]imidazol-2-yl)acetamide 1702-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)-N-(3-ethyl-3,4-D dihydrochromeno[3,4-d]imidazol-2-yl)acetamide 171N-(4H-chromeno[4,3-d]thiazol-2-yl)-2-(1,3-dimethyl-2,6-dioxo-1,2,3,6- Dtetrahydropurin-7-yl)acetamide 172N-(7-chloro-4H-chromeno[4,3-d]thiazol-2-yl)-2-(1,3-dimethyl-2,6- Ddioxo-1,2,3,6-tetrahydropurin-7-yl)acetamide 1732-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydropurin-7-yl)-N-(7-ethoxy- D4H-chromeno[4,3-d]thiazol-2-yl)acetamide

Certain compounds disclosed herein may exist in particular geometric orstereoisomeric forms. The present invention contemplates all suchcompounds, including cis- and trans-isomers, R- and S-enantiomers,diastereomers, (d)-isomers, (l)-isomers, the racemic mixtures thereof,and other mixtures thereof, as falling within the scope of theinvention. For example, if one chiral center is present in a molecule,the invention includes racemic mixtures, enantiomerically enrichedmixtures, and substantially enantiomerically pure compounds. Thecomposition can contain, e.g., more than 50%, more than 60%, more than70%, more than 80%, more than 90%, more than 95%, or more than 99% of asingle enantiomer.

The “enantiomeric excess” or “% enantiomeric excess” of a compositioncan be calculated using the equation shown below. In the example shownbelow a composition contains 90% of one enantiomer, e.g., the Senantiomer, and 10% of the other enantiomer, i.e., the R enantiomer.

ee=(90−10)/100=80%.

Thus, a composition containing 90% of one enantiomer and 10% of theother enantiomer is said to have an enantiomeric excess of 80%.

Methods of preparing substantially isomerically pure compounds are knownin the art. If, for instance, a particular enantiomer of a compounddisclosed herein is desired, it may be prepared by asymmetric synthesis,or by derivation with a chiral auxiliary, where the resultingdiastereomeric mixture is separated and the auxiliary group cleaved toprovide the pure desired enantiomers. Alternatively, where the moleculecontains a basic functional group, such as amino, or an acidicfunctional group, such as carboxyl, diastereomeric salts may be formedwith an appropriate optically active acid or base, followed byresolution of the diastereomers thus formed by fractionalcrystallization or chromatographic means well known in the art, andsubsequent recovery of the pure enantiomers. Alternatively,enantiomerically enriched mixtures and pure enantiomeric compounds canbe prepared by using synthetic intermediates that are enantiomericallypure in combination with reactions that either leave the stereochemistryat a chiral center unchanged or result in its complete inversion.Techniques for inverting or leaving unchanged a particular stereocenter,and those for resolving mixtures of stereoisomers are well known in theart, and it is well within the ability of one of skill in the art tochoose an appropriate method for a particular situation. See, generally,Furniss et al. (eds.), Vogel's Encyclopedia of Practical OrganicChemistry 5^(th) Ed., Longman Scientific and Technical Ltd., Essex,1991, pp. 809-816; and Heller, Acc. Chem. Res. 23: 128 (1990).

The compounds described herein may also contain unnatural proportions ofatomic isotopes at one or more of the atoms that constitute suchcompounds. For example, the compounds may be radiolabeled withradioactive isotopes, such as for example tritium (³H), iodine-125(¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations of the compoundsdisclosed herein, whether radioactive or not, are intended to beencompassed within the scope of the present invention. For example,deuterated compounds and compounds incorporated ¹³C are intended to beencompassed within the scope of the invention.

Certain compounds disclosed herein can exist in unsolvated forms as wellas solvated forms, including hydrated forms. In general, the solvatedforms are equivalent to unsolvated forms and are encompassed within thescope of the present invention. Certain compounds disclosed herein mayexist in multiple crystalline or amorphous forms. In general, allphysical forms are equivalent for the uses contemplated by the presentinvention and are intended to be within the scope of the presentinvention.

As set out above, certain embodiments of the present compounds maycontain a basic functional group, such as amino or alkylamino, and are,thus, capable of forming pharmaceutically acceptable salts withpharmaceutically acceptable acids. The term “pharmaceutically acceptablesalts” in this respect, refers to the relatively non-toxic, inorganicand organic acid addition salts of compounds disclosed herein. Thesesalts can be prepared in situ during the final isolation andpurification of the compounds of the invention, or by separatelyreacting a purified compound of the invention in its free base form witha suitable organic or inorganic acid, and isolating the salt thusformed. Representative salts include the hydrobromide, hydrochloride,sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate,palmitate, stearate, laurate, benzoate, lactate, phosphate, tosylate,citrate, maleate, fumarate, succinate, tartrate, napthylate, mesylate,glucoheptonate, lactobionate, and laurylsulphonate salts and the like.(See, for example, Berge et al. (1977) “Pharmaceutical Salts”, J. Pharm.Sci. 66:1-19.)

In other cases, the compounds disclosed herein may contain one or moreacidic functional groups and, thus, are capable of formingpharmaceutically acceptable salts with pharmaceutically acceptablebases. The term “pharmaceutically acceptable salts” in these instancesrefers to the relatively non-toxic, inorganic and organic base additionsalts of compounds disclosed herein. These salts can likewise beprepared in situ during the final isolation and purification of thecompounds, or by separately reacting the purified compound in its freeacid form with a suitable base, such as the hydroxide, carbonate orbicarbonate of a pharmaceutically acceptable metal cation, with ammonia,or with a pharmaceutically acceptable organic primary, secondary ortertiary amine. Representative alkali or alkaline earth salts includethe lithium, sodium, potassium, calcium, magnesium, and aluminum saltsand the like. Representative organic amines useful for the formation ofbase addition salts include ethylamine, diethylamine, ethylenediamine,ethanolamine, diethanolamine, piperazine and the like. (See, forexample, Berge et al., supra).

An antagonist of TRPA1 function may inhibit the outward current, theinward current, or both currents. Compounds that inhibit both currentsmay do so with the same or with differing IC₅₀ values. The inhibition ofa particular current is measured by the ability to inhibit or reducesuch current (e.g., inward and/or outward) in an in vitro or an in vivoassay. Compounds that inhibit any of the foregoing currents in an invitro or in vivo assay are characterized as compounds that inhibit afunction of TRPA1. Additionally or alternatively, a further exemplaryfunction of TRPA1 that may be inhibited by the present compounds is ionflux mediated by TRPA1.

A compound described herein (e.g., a TRPA1 antagonist) can be chosen fora use described herein may be chosen because it inhibits a TRPA1function with an IC₅₀ less than or equal to 10 uM, 5 uM, 1 uM, or lessthan or equal to 700, 600, 500, 400, 300, 250, 200, or 100 nM. In otherembodiments, the compound described herein inhibits a TRPA1 functionwith an IC₅₀ less than or equal to 75 nM, less than or equal to 50 nM,or less than or equal to 25, 10, 5, or 1 nM.

In certain embodiments, inhibition of TRPA1 function means that afunction, for example a TRPA1 mediated current, is decreased by greaterthan 50% in the presence of an effective amount of a compound incomparison to in the absence of the compound or in comparison to anineffective amount of a compound. In certain other embodiments, theinhibition of a TRPA1 function means that a function, for example aTRPA1 mediated current or TRPA1 mediated ion flux, is decreased by atleast 50%, 60%, 70%, 75%, 80%, 85%, or 90% in the presence of aneffective amount of a compound in comparison to in the absence of thecompound. In still other embodiments, the inhibition of a TRPA1 functionmeans that a function, for example a TRPA1 mediated current, isdecreased by at least 92%, 95%, 97%, 98%, 99%, or 100% in the presenceof an effective amount of a compound in comparison to in the absence ofthe compound.

In some embodiments, a compound described herein (e.g., a TRPA1antagonist) can be characterized by some level of activity versus otherion channels (e.g., certain compounds are selective for inhibiting TRPA1and other compounds exhibit a level of cross reactivity against one ormore other ion channel). When a compound described herein ischaracterized by its activity against another ion channel, inhibition ofa function or activity of the other ion channel is defined analogouslyto the way in which a function of a TRPA1 channel is defined. Thus,inhibiting the function of another ion channel means, for example,inhibiting ion flux mediated by that other ion channel or inhibiting thecurrent mediated by that other ion channel.

In some embodiments, a compound described herein (e.g., a TRPA1antagonist) is chosen for use because it is more selective for one TRPisoform than others, e.g., 10-fold, and more preferably at least 20, 40,50, 60, 70, 80, or at least 100- or 1000-fold more selective for TRPA1over one or more of TRPC6, TRPV5, TRPV6, TRPM8, TRPV1, TRPV2, TRPV4,and/or TRPV3. In other embodiments, the differential is smaller, e.g.,it more strongly inhibits TRPA1 than TRPM8, TRPV1, TRPV2, TRPV3, and/orTRPV4, preferably at least twice, three times, five times, or ten timesmore strongly. Such comparisons may be made, for example, by comparingIC₅₀ values.

In other embodiments, a compound described herein (e.g., a TRPA1antagonist) is chosen for use because it is more selective for TRPA1than for other non-TRP ion channels, e.g., 10-fold, and more preferablyat least 20, 40, 50, 60, 70, 80, or at least 100- or 1000-fold moreselective for TRPA1 over one or more of NaV1.2, Cav1.2, Cav3.1, HERG,and/or mitochondrial uniporter. In other embodiments, the differentialis smaller, e.g., it more strongly inhibits TRPA1 than NaV1.2, Cav1.2,Cav3.1, HERG, and/or mitochondrial uniporter, preferably at least twice,three times, five times, or ten times more strongly.

In certain embodiments, the compound described herein (e.g., a TRPA1antagonist) inhibits TRPA1 with an IC₅₀ at least one order of magnitudemore potent than its Ki for thealpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)receptor. In certain other embodiments, the compound described herein(e.g., a TRPA1 antagonist) inhibits TRPA1 with an IC₅₀ at least twoorders of magnitude, three orders of magnitude, or four orders ofmagnitude more potent than its Ki for the AMPA receptor. In certainembodiments, the compound described herein (e.g., a TRPA1 antagonist)does not appreciably bind the AMPA receptor. In other words, the subjectantagonists inhibit TRPA1 with a particular IC₅₀ and, when administeredat that concentration, the antagonist does not appreciably bind the AMPAreceptor.

In certain embodiment, a compound described herein is chosen because itantagonizes the function of TRPA1 and the function of TRPM8, TRPV1and/or TRPV3. Although such compounds selectively antagonize thefunction of more than one ion channel, the IC₅₀ values for the differention channels need not be identical.

The IC₅₀ values are measured in vitro using, for example, patch clampanalysis or standard measurements of calcium flux. Exemplary in vitromethods for calcium flux-based IC₅₀ estimation are described in Example2. Methods used to obtain more definitive IC₅₀ measurements aredescribed in Example 3. Alternatively, estimates of % inhibition ofcurrent or ion flux can also be calculated and used to assess efficacyof a compound as an inhibitor.

Indications

Movement of ions across cellular membranes is carried out by specializedproteins. TRP channels are one large family of non-selective cationchannels that function to help regulate ion flux and membrane potential.Non-selective cation channels such as TRPA1 modulate the flux of calciumand sodium ions across cellular membranes. Sodium and calcium influxleads to depolarization of the cell. This increases the probability thatvoltage-gated ion channels will reach the threshold required foractivation. As a result, activation of non-selective cation channels canincrease electrical excitability and increase the frequency ofvoltage-dependent events. Voltage-dependent events include, but are notlimited to, neuronal action potentials, cardiac action potentials,smooth muscle contraction, cardiac muscle contraction, and skeletalmuscle contraction.

Calcium influx caused by the activation of non-selective cation channelssuch as TRPA1 also alters the intracellular free calcium concentration.Calcium is a ubiquitous second messenger molecule within the cell, soalterations in intracellular calcium levels have profound effects onsignal transduction and gene expression. As a result, activation ofnon-selective cation channels such as TRPA1 can lead to changes in geneexpression and cellular phenotype. Gene expression events include, butare not limited to, production of mRNAs encoding cell surface receptors,ion channels, and kinases. These changes in gene expression can lead tohyperexcitability in that cell.

Modulating the function of TRPA1 proteins provides a means of modulatingcalcium homeostasis, sodium homeostasis, membrane polarization, and/orintracellular calcium levels, and compounds that can modulate TRPA1function are useful in many aspects, including, but not limited to,maintaining calcium homeostasis, modulating intracellular calciumlevels, modulating membrane polarization, and treating or preventingdiseases, disorders, or conditions associated with calcium and/or sodiumhomeostasis or dyshomeostasis.

Bautista et al. have reported that TRPA1 mediates the inflammatoryactions of agents such as acrolein, which causes the inflammatory andtoxic actions of tear gas. (Bautista et al., Cell (2006) 124:1269-82.)Bessac et al. have shown that hypochlorite—the main mediator of chlorineirritancy—activates Ca²⁺ influx and membrane currents in sensory neuronscultured from Trpa1^(+/+) mice, while the influx of Ca²⁺ was absent inneurons cultured from Trpa1^(−/−) mice. In addition, Bessac et al.compared the responses of Trpa1^(+/+) mice and Trpa1^(−/−) mice to NaOClexposure. The Trpa1^(+/+) mice showed much more respiratory depressionthan did the Trpa1^(−/−) mice. (Bessac et al., Journal of ClinicalInvestigation (2008) 18:1899-1910) These data suggest that TRPA1antagonists could be useful to treat or prevent the effects of chemicalwarfare agents including tear gas and chlorine.

Chemical Warfare Agents

A subject may be exposed to a chemical warfare agent, e.g., byinhalation or by contact with the skin. If a compound described herein(e.g., a TRPA1 antagonist) is administered, the symptoms or injuriesresulting from the exposure to the chemical warfare agents can bereduced, prevented, or both. The compound described herein can beadministered to a subject before, during, or following such exposure andis therefore administered within 24 hours, 18 hours, 6 hours, 4 hours, 3hours, 2 hours, 1 hour, 30 minutes, 20 minutes, 10 minutes, 5 minutes,one minute, or thirty seconds before or after such exposure. Thecompound described herein can be administered prophylactically, whenexposure to an agent is anticipated. It can also be administered afterexposure to the chemical warfare agent agent (e.g., before or aftersymptoms of injury present in a subject).

Injuries resulting from the exposure to chemical warfare agents areknown in the art and include any physical injuries, such as injuries tothe skin, eyes, respiratory tract, musculo-skeletal system, circulatorysystem, gastrointestinal tract, central nervous system, peripheralnervous system, heart, liver, lungs, and kidneys. Exemplary symptoms orinjuries resulting from the exposure to chemical warfare agents includeinflammation, burn, itch, pain, rash, blisters, sweating, muscletwitching, nausea, vomiting, diarrhea, weakness, loss of conciousness,convulsions, muscular twitching, paralysis, secretions (from the mouth,nose, or lung for example), difficulty breating, blurred vision, eyepain, lacrimation, red eyes, shortness of breath, coughing, phlegmproduction and narrowing of the airways, headaches, tremors, dizziness,numbness or tingling, anxiety, insomnia, depression, emotionalinstability, and even death. The term “chemical warfare agent” includesall of those agents classified as schedule 1, 2, and 3 agents under theChemical Weapons Convention of 1993 and may be in liquid form, gas form,solid form, or combinations thereof. Exemplary agents are described infurther detail below and include, for example, nerve agents, bloodagents, blister agents, pulmonary agents, incapacitating agents, andtoxins. Other agents include methyl isocyanate, hexamethylenediisocyanate, 2,4-toluene-diisocyanate, anddiphenylmethane-4,4-diisocyanate (MDI).

Nerve agents. Nerve agent poisoning typically leads to contraction ofpupils, profuse salivation, convulsions, involuntary urination anddefecation, and eventual death by asphyxiation as control is lost overrespiratory muscles. These symptoms are reduced or prevented by theadministration of the TRPA1 antagonists. Exemplary agents include Gagents such as tabun (GA), sarin (GB), soman (GD), cyclosarin (GF), andGV; V agents such as VE, VG, VM, VX, and Novichok agents.

Blood agents. A blood agent (or cyanogen agent) is a compound containinga cyanide group that prevents the body from utilizing oxygen. Theseagents exert their toxic effect at the cellular level by directlyinterrupting cellular respiration. Exemplary agents include cyanogenchloride, hydrogen cyanide, and hydrogen sulfide.

Blister agents. Blister agents or vesicants typically cause severe skin,eye and mucosal pain and irritation. These agents also have the abilityto cause large, painful water blisters. Blister agents include, forexample, lewisites, nitrogen mustard, sulfur mustard,ethyldichloroarsine (a lewisite analog; ED), methyldichloroarsine (MD),phenyldichloroarsine (PD), and phosgene oxime (CX). Lewisites include,for example, 2-Chlorovinyldichloroarsine (Lewisite 1),Bis(2-chlorovinyl)chloroarsine (Lewisite 2), andTris(2-chlorovinyl)arsine (Lewisite 3). Exemplary nitrogen mustards arebis(2-chloroethyl)ethylamine (HN1), bis(2-chloroethyl)methylamine (HN2),and tris(2-chloroethyl)amine (HN3). Sulfur mustards include, forexample, 1,2-Bis(2-chloroethylthio) ethane (Sesquimustard; Q),1,3-Bis(2-chloroethylthio)-n-propane,1,4-Bis(2-chloroethylthio)-n-butane,1,5-Bis(2-chloroethylthio)-n-pentane, 2-Chloroethylchloromethylsulfide,bis(2-chloroethyl) sulfide (Mustard gas; HD), bis(2-chloroethylthio)methane, bis(2-chloroethylthiomethyl) ether, andbis(2-chloroethylthioethyl) ether (O Mustard).

Pulmonary agents. A pulmonary agent (or choking agent) is a chemicalweapon agent designed to impede a subject's ability to breathe,resulting in suffocation. Exemplary agents include adamsite (DM),acrolein, bis(chloromethyl) ether (BCME), chlorine (Cl₂), chloropicrin(PS), diphosgene (DP), methyl chlorosulfonate, phosgene (CG), andstannic chloride.

Incapacitating agents. Incapacitating agents or riot-control agentstypically produce temporary physiological or mental effects, or both,such that individuals who are exposed to them are incapable of concertedeffort. Upon their exposure, lachrymatory agents (or lachrymators) forexample, irritate the eyes to cause tearing, pain, and even temporaryblindness. The most common lachrymatory agents are tear gas and pepperspray and include, for example, a-Chlorotoluene, benzyl bromide,bromoacetone (BA), bromobenzylcyanide (CA) bromomethyl ethyl ketone,capsaicin (OC), chloracetophenone (Tear gas; CN), chloromethylchloroformate, dibenzoxazepine (CR), ethyl iodoacetate,ortho-chlorobenzylidene malononitrile (Super tear gas; CS),trichloromethyl chloroformate, and xylyl bromide. Other incapacitatingagents include, for example, 3-Quinuclidinyl benzilate (psychedelic;BZ), hydrocyanic acid (paralytic), diphenylchloroarsine (sternutatory;DA), diphenylcyanoarsine (DC), and KOLOKOL-1 (tranquilizer).

Toxins. Exemplary toxins are abrin, ricin, and saxitoxin.

Pharmaceutical Compositions

While it is possible for a compound disclosed herein to be administeredalone, it is preferable to administer the compound as a pharmaceuticalformulation, where the compound is combined with one or morepharmaceutically acceptable excipients or carriers. The compoundsdisclosed herein may be formulated for administration in any convenientway for use in human or veterinary medicine. In certain embodiments, thecompound included in the pharmaceutical preparation may be activeitself, or may be a prodrug, e.g., capable of being converted to anactive compound in a physiological setting.

The phrase “pharmaceutically acceptable” is employed herein to refer tothose compounds, materials, compositions, and/or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of human beings and animals without excessive toxicity,irritation, allergic response, or other problem or complication,commensurate with a reasonable benefit/risk ratio.

Examples of pharmaceutically acceptable carriers include: (1) sugars,such as lactose, glucose and sucrose; (2) starches, such as corn starchand potato starch; (3) cellulose, and its derivatives, such as sodiumcarboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4)powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients,such as cocoa butter and suppository waxes; (9) oils, such as peanutoil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil andsoybean oil; (10) glycols, such as propylene glycol; (11) polyols, suchas glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters,such as ethyl oleate and ethyl laurate; (13) agar; (14) bufferingagents, such as magnesium hydroxide and aluminum hydroxide; (15) alginicacid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer'ssolution; (19) ethyl alcohol; (20) phosphate buffer solutions; (21)cyclodextrins such as Captisol®; and (22) other non-toxic compatiblesubstances employed in pharmaceutical formulations.

Examples of pharmaceutically acceptable antioxidants include: (1) watersoluble antioxidants, such as ascorbic acid, cysteine hydrochloride,sodium bisulfate, sodium metabisulfite, sodium sulfite and the like; (2)oil-soluble antioxidants, such as ascorbyl palmitate, butylatedhydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propylgallate, alpha-tocopherol, and the like; and (3) metal chelating agents,such as citric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol,tartaric acid, phosphoric acid, and the like.

Solid dosage forms (e.g., capsules, tablets, pills, dragees, powders,granules and the like) can include one or more pharmaceuticallyacceptable carriers, such as sodium citrate or dicalcium phosphate,and/or any of the following: (1) fillers or extenders, such as starches,lactose, sucrose, glucose, mannitol, and/or silicic acid; (2) binders,such as, for example, carboxymethylcellulose, alginates, gelatin,polyvinyl pyrrolidone, sucrose and/or acacia; (3) humectants, such asglycerol; (4) disintegrating agents, such as agar-agar, calciumcarbonate, potato or tapioca starch, alginic acid, certain silicates,and sodium carbonate; (5) solution retarding agents, such as paraffin;(6) absorption accelerators, such as quaternary ammonium compounds; (7)wetting agents, such as, for example, cetyl alcohol and glycerolmonostearate; (8) absorbents, such as kaolin and bentonite clay; (9)lubricants, such a talc, calcium stearate, magnesium stearate, solidpolyethylene glycols, sodium lauryl sulfate, and mixtures thereof; and(10) coloring agents.

Liquid dosage forms can include pharmaceutically acceptable emulsions,microemulsions, solutions, suspensions, syrups and elixirs. In additionto the active ingredient, the liquid dosage forms may contain inertdiluents commonly used in the art, such as, for example, water or othersolvents, solubilizing agents and emulsifiers, such as ethyl alcohol,isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol,benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (inparticular, cottonseed, groundnut, corn, germ, olive, castor and sesameoils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fattyacid esters of sorbitan, and mixtures thereof.

Suspensions, in addition to the active compounds, may contain suspendingagents as, for example, ethoxylated isostearyl alcohols, polyoxyethylenesorbitol and sorbitan esters, microcrystalline cellulose, aluminummetahydroxide, bentonite, agar-agar and tragacanth, and mixturesthereof.

Ointments, pastes, creams and gels may contain, in addition to an activecompound, excipients, such as animal and vegetable fats, oils, waxes,paraffins, starch, tragacanth, cellulose derivatives, polyethyleneglycols, silicones, bentonites, silicic acid, talc and zinc oxide, ormixtures thereof.

Powders and sprays can contain, in addition to an active compound,excipients such as lactose, talc, silicic acid, aluminum hydroxide,calcium silicates and polyamide powder, or mixtures of these substances.Sprays can additionally contain customary propellants, such aschlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, suchas butane and propane.

The formulations may conveniently be presented in unit dosage form andmay be prepared by any methods well known in the art of pharmacy. Theamount of active ingredient which can be combined with a carriermaterial to produce a single dosage form will vary depending upon thehost being treated, the particular mode of administration. The amount ofactive ingredient that can be combined with a carrier material toproduce a single dosage form will generally be that amount of thecompound which produces a therapeutic effect. Generally, out of onehundred percent, this amount will range from about 1 percent to aboutninety-nine percent of active ingredient, preferably from about 5percent to about 70 percent, most preferably from about 10 percent toabout 30 percent.

The tablets, and other solid dosage forms of the pharmaceuticalcompositions disclosed herein, such as dragees, capsules, pills andgranules, may optionally be scored or prepared with coatings and shells,such as enteric coatings and other coatings well known in thepharmaceutical-formulating art. They may also be formulated so as toprovide slow or controlled release of the active ingredient thereinusing, for example, hydroxypropylmethyl cellulose in varying proportionsto provide the desired release profile, other polymer matrices,liposomes and/or microspheres. They may be sterilized by, for example,filtration through a bacteria-retaining filter, or by incorporatingsterilizing agents in the form of sterile solid compositions that can bedissolved in sterile water, or some other sterile injectable mediumimmediately before use. These compositions may also optionally containopacifying agents and may be of a composition that they release theactive ingredient(s) only, or preferentially, in a certain portion ofthe gastrointestinal tract, optionally, in a delayed manner Examples ofembedding compositions that can be used include polymeric substances andwaxes. The active ingredient can also be in micro-encapsulated form, ifappropriate, with one or more of the above-described excipients.

Dosage forms for the topical or transdermal administration of a compoundof this invention include powders, sprays, ointments, pastes, creams,lotions, gels, solutions, patches and inhalants. The active compound maybe mixed under sterile conditions with a pharmaceutically acceptablecarrier, and with any preservatives, buffers, or propellants that may berequired.

The formulations disclosed herein can be delivered via a device.Exemplary devices include, but are not limited to, a catheter, wire,stent, or other intraluminal device. Further exemplary delivery devicesalso include a patch, bandage, mouthguard, or dental apparatus.Transdermal patches have the added advantage of providing controlleddelivery of a compound disclosed herein to the body. Such dosage formscan be made by dissolving or dispersing the compound in the propermedium. Absorption enhancers can also be used to increase the flux ofthe compound across the skin. The rate of such flux can be controlled byeither providing a rate controlling membrane or dispersing the compoundin a polymer matrix or gel.

Another example of a device is a metered dose aerosol dispensercontaining an aerosol pharmaceutical composition for pulmonary or nasaldelivery comprising an agent that inhibits a TRPA1-mediated current withan IC₅₀ of 1 micromolar or less. For instance, it can be a metered doseinhaler, a dry powder inhaler or an air-jet nebulizer.

Ophthalmic formulations, eye ointments, drops, solutions and the like,are also contemplated as being within the scope of this invention.

In some cases, in order to prolong the effect of a drug, it is desirableto slow the absorption of the drug from subcutaneous or intramuscularinjection. This may be accomplished by the use of a liquid suspension ofcrystalline or amorphous material having poor water solubility. The rateof absorption of the drug then depends upon its rate of dissolution,which, in turn, may depend upon crystal size and crystalline form.Alternatively, delayed absorption of a parenterally administered drugform is accomplished by dissolving or suspending the drug in an oilvehicle.

Injectable depot forms are made by forming microencapsule matrices ofthe subject compounds in biodegradable polymers such aspolylactide-polyglycolide. Depending on the ratio of drug to polymer,and the nature of the particular polymer employed, the rate of drugrelease can be controlled. Examples of other biodegradable polymersinclude poly(orthoesters) and poly(anhydrides). Depot injectableformulations are also prepared by entrapping the drug in liposomes ormicroemulsions that are compatible with body tissue.

When the compounds disclosed herein are administered as pharmaceuticals,to humans and animals, they can be given per se or as a pharmaceuticalcomposition containing, for example, 0.1 to 99.5% (more preferably, 0.5to 90%) of active ingredient in combination with a pharmaceuticallyacceptable carrier.

The formulations can be administered topically, orally, transdermally,rectally, vaginally, parentally, intranasally, intrapulmonary,intraocularly, intravenously, intramuscularly, intraarterially,intrathecally, intracapsularly, intraorbitally, intracardiacly,intradermally, intraperitoneally, transtracheally, subcutaneously,subcuticularly, intraarticularly, subcapsularly, subarachnoidly,intraspinally, intrasternally or by inhalation.

Particularly useful modes of administration include topicaladministration, intramuscular injection, inhalation, topical ocularadministration (e.g., via eye drops), or oral administration.

Combination Therapy

Another aspect of the invention provides a conjoint therapy wherein oneor more other therapeutic agents are administered with a compounddescribed herein (e.g., a TRPA1 modulator such as an antagonist). Suchconjoint treatment may be achieved by way of the simultaneous,sequential, or separate dosing of the individual components of thetreatment.

In certain embodiments, two or more compounds of the invention areconjointly administered. When two or more compounds of the invention areconjointly administered, the two or more compounds may have a similarselectivity profile and functional activity, or the two or morecompounds may have a different selectivity profile and functionalactivity. By way of example, the two or more compounds may both beapproximately 10, 100, or 1000 fold selective for antagonizing afunction of TRPA1 over TRPV1, TRPV5, and TRPV6 (e.g., the two or morecompounds have a similar selectivity profile), and further may inhibit afunction of TRPA1 with a similar IC50 (e.g., a similar functionalactivity). Alternatively, the one of the two or more compounds mayselectively inhibit TRPA1 while the other of the two or more compoundsinhibits both TRPA1 and TRPV1 (e.g., the two or more compounds havediffering selectivity profiles). Administration of combinations of twoor more compounds of the invention having similar or differingproperties are contemplated.

In certain embodiments, a compound of the invention is conjointlyadministered with one or more additional compounds that antagonize thefunction of a different channel. By way of example, a compound of theinvention may be conjointly administered with one or more compounds thatantagonize TRPV1, TRPM8, and/or TRPV3. The compound(s) that antagonizeTRPV1, TPRM8, or TRPV3 may be selective for TRPV1, TRPM8 or TRPV3 (e.g.,inhibit TRPV1 or TRPV3 10, 100, or 1000 fold more strongly than TRPA1).Alternatively, the compound(s) that antagonize TRPV1 or TRPV3 may crossreact with other TRP channels.

In certain other embodiments, a compound of the invention is conjointlyadministered with one or more additional agents or therapeutic regimensappropriate for the particular injury being treated. For example,current treatments for injuries caused by exposure to nerve agentsinclude treatment with atropine and Pralidoxime chloride (2-PAM).Injuries caused by exposure to cyanides are currently treated with.Scavengers such as human plasma-derived butyrylcholinesterase (HuBuChE),anti-epilectic drugs and neutralizing decontamination solutions such as0.5% hypochloriate are also used to treat such injuries. Treatments forcyanide poisoning include hydroxocobalamin, sodium nitrite, andrhodanase. M291 resin kits are also currently used to treat topicalinjuries resulting from exposure to chemical warfare agents. Any ofthese agents can be combined with the TRPA1 antagonists describedherein.

Dosages

Actual dosage levels of the active ingredients in the pharmaceuticalcompositions of this invention may be varied so as to obtain an amountof the active ingredient that is effective to achieve the desiredtherapeutic response for a particular patient, composition, and mode ofadministration, without being toxic to the patient.

The selected dosage level will depend upon a variety of factorsincluding the activity of the particular compound disclosed hereinemployed, or the ester, salt or amide thereof, the route ofadministration, the time of administration, the rate of excretion of theparticular compound being employed, the duration of the treatment, otherdrugs, compounds and/or materials used in combination with theparticular compound employed, the age, sex, weight, condition, generalhealth and prior medical history of the patient being treated, and likefactors well known in the medical arts.

A physician or veterinarian having ordinary skill in the art can readilydetermine and prescribe the effective amount of the pharmaceuticalcomposition required. For example, the physician or veterinarian couldstart doses of the compounds of the invention employed in thepharmaceutical composition at levels lower than that required in orderto achieve the desired therapeutic effect and gradually increase thedosage until the desired effect is achieved.

In general, a suitable daily dose of a compound of the invention will bethat amount of the compound that is the lowest dose effective to producea therapeutic effect. Such an effective dose will generally depend uponthe factors described above. Generally, intravenous,intracerebroventricular and subcutaneous doses of the compounds of thisinvention for a patient will range from about 0.0001 to about 100 mg perkilogram of body weight per day. For example, the dose can be 1-50,1-25, or 5-10 mg/kg.

Disease and Injury Models

Compounds that antagonize TRPA1 function may be useful in theprophylaxis and treatment of any of the foregoing injuries, diseases,disorders, or conditions. In addition to in vitro assays of the activityof these compounds, their efficacy can be readily tested in one or moreanimal models. Compounds that may reduce pain or other undesirablesymptoms in the animals can be readily tested by observing behavioraland/or physical characteristics of challenged animals in the presenceversus the absence of the test compound(s) or procedure.

Mouse Ear Edema Model. The mouse ear edema model provides a quantitativeedema response as well as histopathological and biochemical endpoints asmeasurements of inflammation and tissue damage following exposure to thechemical warfare agent half mustard. The model is performed using CD-1mice. Twenty-four hours following IP administration of test compound orvehicle, a solution of half mustard in organic solvent is applied to theinner surface of the right ear of each mouse. The left ear is nottreated with half mustard and instead is maintained as a control.Animals are sacrificed at 24 hours, 72 hours, and 7 days post exposureto half mustard. The ear weights, draize scoring, and pathology arerecorded at each time point. Half mustard injury in the mouse ear ismeasured by both edema response (fluid accumulation) andhistopathological damage (necrosis, epidermal-dermal separation).

In an alternate model, the test compound is administeredtherapeutically, e.g., immediately after, or one hour after exposure tohalf mustard. The models can also be performed using sulfur mustard ornitrogen mustard instead of half mustard.

Guinea Pig Model for Nerve Agent Challenge. A Guinea pig model can beused to study the protective effects of the compounds against mortalityinduced by a nerve agent. The animals are pretreated with two IP dosesof test compound, one IM dose of atropine, one IM dose of 2-PAM, or twoIP doses of vehicle before challenge with 2×LD50 (s.c.) of a nerveagent. The animals are observed at 6 observation times post challenge.The number of animals in each group surviving 24 hours post challenge isthen recorded.

In an alternate model, the test compound, atropine, and 2-PAM areadministered therapeutically, e.g., immediately after, or one hour afterexposure to the nerve agent.

Any of the foregoing animal models may be used to evaluate the efficacyof a TRPA1 inhibitor in treating pain associated with pancreatitis. Theefficacy can be compared to a no treatment or placebo control.Additionally or alternatively, efficacy can be evaluated in comparisonto one or more known pain relieving medicaments.

The following examples are meant to be illustrative and are not meant tobe limiting in any way.

Example 1: Synthesis of Exemplary Compounds General Procedure A for thePreparation of Amides by Coupling Using EDCI

To a mixture of theophylline-7-acetic acid (2 mmol), DMAP (2 mmol),substituted phenethylamine (2 mmol) and DIPEA (4 mmol) in DMF (20 mL) isadded EDCI (2 mmol). The reaction mixture is heated to 40° C. andstirred over night. The solution is concentrated in vacuo and theresidue is dissolved in EtOAc (100 mL), washed with H₂O, citric acid(10%), NaHCO₃ (sat.) and brine, dried over Na₂SO₄ and concentrated invacuo. The crude product is purified by flash chromatography on silicagel eluting with MeOH/EtOAc (1-8%).

General Procedure B for the Preparation of Amides Via Acid Chloride

A suspension of theophylline-7-acetic acid (2 mmol) in CHCl₃ (15 mL) andMeCN (15 mL) is cooled in an ice-water bath. Oxalyl chloride (2.2 mmol)is then added dropwise. Catalytic DMF (˜25 μL) is then added. Themixture is stirred at room temperature over night. The solution is thencooled in an ice-water bath, and DMAP (2.5 mmol) is added in oneportion. The substituted phenethylamine is added dropwise and thereaction mixture is stirred at room temperature over night. Afterdiluting with CHCl₃ (50 mL), the mixture is washed with H₂O, citric acid(10% in H₂O), NaHCO₃ (sat.), dried over Na₂SO₄ and concentrated invacuo. The crude product is purified by flash chromatography on silicagel eluting with MeOH/EtOAc (1˜8%).

As shown in Scheme 1, dihydropyrimidine-dione 2 can be prepared byreacting 1-propylurea (1) and ethyl 2-cyanoacetate, which can besubsequently treated with bromine, ethyl 2-aminoacetate, andtriethoxymethane to yield compound 6, ethyl2-(2,6-dioxo-1-propyl-2,3-dihydro-1H-purin-7(6H)-yeacetate. The obtaineddihydropurine 6 can be transformed to compound 10 through methylation,hydrolysis, and a coupling reaction under CDI.

As shown in Scheme 2, dihydropyrimidine-dione 2 can be prepared byreacting 1-propylurea (1) and ethyl 2-cyanoacetate, which can besubsequently treated with bromine, ethyl 2-aminoacetate, andtriethoxymethane to yield compound 6, ethyl2-(1-methyl-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate. Theobtained dihydropurine 6 can be converted to compound 10 throughalkylation reaction, hydrolysis, and a coupling reaction catalyzed byCDI.

As shown in Scheme 3, N,N-Dimethylethane-1,2-diamine can be converted tourea 2, which can then react with ethyl 2-cyanoacetate to givedihydropyrimidine-dione 3. Compound 3 can be subsequently treated withbromine, ethyl 2-aminoacetate, and triethoxymethane to yield compound 7,ethyl2-(1-(2-(dimethylamino)ethyl)-2,6-dioxo-2,3-dihydro-1H-purin-7(6H)-yl)acetate.The obtained dihydropurine 7 can be transformed to compound 11 throughalkylation reaction, hydrolysis, and a coupling reaction catalyzed byCDI.

As shown in Scheme 4, esterification of dihydropurine 1, followed byreduction with LAH, Swern oxidation and coupling reaction can yieldcompound 5, which subsequently can be converted to compound 6, compound7, and compound 8 through methylation, acylation, or sulphonylation.

As shown in Scheme 5, dihydropurine 1 can be coupled with2-(4-methylpiperazin-1-yl)ethanamine by CDI to give compound 2.

As shown in Scheme 6, 2-(4-Phenylpiperazin-1-yl)ethanamine 4 can beprepared by reacting 1-phenylpiperazine with 2-chloroacetamide, followedby a reduction reaction with LAH. Amine 4 then can be coupled withdihydropurine 5 to yield compound 6.

As shown in Scheme 7, 2-(1-Benzyl-1H-imidazol-2-yl)ethanamine 4 can beprepared by protection of imidazole, followed by alkylation and adeprotection reaction with TFA. Amine 4 then can be coupled withdihydropurine 5 to afford compound 6.

As shown in Scheme 8, treatment of imidazole 1 with n-BuLi, followed byan alkylation reaction, and a deprotection reaction with TFA affords2-(1-methyl-1H-imidazol-2-yl)ethanamine. Amine 3 can be coupled withdihydropurine 4 to give compound 5.

As shown in Scheme 9, 2-(Thiazol-2-yl)ethanamine 3 can be prepared bytreatment of thiazole 1 with n-BuLi, followed by addition ofBoc-protected 2-bromoethanamine and a deprotection reaction with TFA.The obtained amine 3 then can be coupled with carboxylic acid 4 toafford compound 5.

As shown in Scheme 11, treatment of oxazole 1 with n-BuLi, followed byan alkylation reaction, and a deprotection reaction with TFA affords2-(oxazol-2-yl)ethanamine 4 Amine 4 can be coupled with dihydropurine 5to give compound 6.

Compound 10 (Scheme 12) can be prepared according to similar reactionprocedures shown in Scheme 1.

As shown in Scheme 13, treatment of Protected indoline 2 with LDA anddibromopropane gives compound 3, which subsequently can react withpurine-dione 4, followed by hydrolysis reaction to yield compound 6.

As shown in Scheme 14, 2-(5-Methylpyridin-2-yl)ethanamine 4 can beprepared by converting 2-chloro-5-methylpyridine (1) to2-bromo-5-methylpyridine, followed by reacting with Boc-protected2-bromoethanamine and removal of the protecting group with TFA. Theobtained amine 4 then can be coupled with carboxylic acid 5 to affordcompound 6.

As shown in Scheme 16, LAH can reduce ethyl 6-methylnicotinate to givealcohol 2, which can be oxidized and subsequently treated with MeNO₂ toyield compound 4. Compound 4 can be reduced to amine 4, which can becoupled with carboxylic acid 6 to give compound 7.

Additional compounds can be synthesized as described in WO 2007/073505.

Example 2: High Thoughput Screening Assay

The assay depended on detection of the rise in intracellular Ca²⁺concentration ([Ca²⁺]_(i)) following channel activation in cellsinducibly expressing the TRPA1 channel. Ca²⁺ rise was quantified withthe use of fluorescent Ca²⁺ indicators that were loaded into cells andthereafter indicated the [Ca²⁺]_(i). Ca²⁺ influx followed activation ofthe TRPA1 channel Compounds inhibiting the [Ca²⁺]_(i) rise wereconsidered hits for further investigation.

The commercially available HEK293/TREx line (Invitrogen) was stablytransfected with a TRPA1 construct (specifically a construct encoding aTRPA1 protein with an amino acid sequence depicted in SEQ ID NO: 1 of WO2007/073505) and screened by conventional calcium imaging to find cloneswith TRPA1 expression following stimulation with 1 μg/ml tetracycline.These cells were maintained in the growth medium recommended by themanufacturer supplemented with 100 μg/ml hygromycin to promote retentionof the TRPA1 construct. After growing to near confluency, cells wereplated at a density of ˜25,000 cells/well in 384 well CellBind plates(Corning) in the presence of 1 μg/ml tetracycline, and allowed to growfor 20-30 hrs. A nearly confluent monolayer resulted. Cells were thenloaded with Ca²⁺ dye: Fura-2/AM or Fluo4/AM was added to the wells to afinal concentration of 2 μM or 1 μM, respectively, and incubated for ˜60min at room temperature. Supernatant was then removed from the cells byinverting plates with a sharp flick, and Hank's Balanced Salt Solution(HBSS; 0.185 g/l D-glucose, 0.9767 g/l MgSO₄ (anhydrous), 0.4 g/l KCl,0.06 g/l KH₂PO₄ (anhydrous), 0.35 g/l NaHCO₃, 8.0 g/l NaCl, and 0.04788g/l Na₂HPO₄ (anhydrous); pH 7.4) was then added to each well. Followingrecovery from loading, cells were assayed using the Hamamatsu FDSS 6000system, which permitted illumination alternately at 340 nM and 380 nMfor Fura-2 experiments, or at 485 nM for Fluo4 experiments. Frames wereacquired at a rate of 0.2 Hz. For the screening assay, a diluted stock(at 50 μM) of compounds to be tested was added to each well for 2minutes following the collection of a short (4 frame) baseline. AITC(allylisothiocyanate) was then added to each well, achieving a finalconcentration of 10 μM each compound and 7.5 μM AITC. Data werecollected for at least 3 minutes following addition of AITC, andevaluated for the [Ca²⁺]_(i), which is proportional to the fluorescentintensity (for Fluo4) or the F340/F380 ratio (for Fura-2). Negativecontrols consisted of HEK293/TREx TRPA1 cells exposed to AITC, but nocompound. Positive control cells were usually HEK293/TREx (“parental”)cells exposed to AITC but no compound, but sometimes normal HEK/293 TRExTRPA1 cells were also used, but not exposed to AITC or compound. Thesecontrols defined a screening window, and “hits” were defined as thosecompounds inhibiting the fluorescence response by at least 40%. IC₅₀values were determined for compounds defined as “hits.” The Fluo4cell-based fluorescence assay was used to determine the intracellularCa²⁺ concentration in the presence of varying drug concentration. Todetermine IC₅₀ values, concentrations tested were 40 μM, 20 μM, 10 μM, 5μM, 2.5 μM, 1.25 μM, and 0.625 μM. Compounds were tested in triplicateat all concentrations. Standard software was used to fit IC₅₀ curves.

Additionally or alternatively, potency can be represented as %inhibition of a response in the presence (of a given concentration ofcompound) versus the absence of compound or in comparison to a controlcompound. For example, efficacy can be represented as % inhibition ofion flux in the presence versus the absence of compound.

Example 3: Patch Clamp Experiments

Patch clamp experiments permit the detection of currents through theTRPA1 channel in the cell line described above. The whole-cellconfiguration of the patch clamp technique was used to test thecompounds described herein. In normal whole-cell patch clamp recordings,a glass electrode is brought into contact with a single cell and ahigh-resistance (gigaohm) seal is established with the cell membrane.The membrane is then ruptured to achieve the whole-cell configuration,permitting control of the voltage of the cell membrane and measurementof currents flowing across the membrane using the amplifier attached tothe electrode and resulting in the replacement of cytoplasm with thepipette solution.

HEK293/TREx TRPA1 cells were induced in the presence of 1 μg/mltetracycline for 20-48 hours, removed from growth plates, and replatedat low density (to attain good single-cell physical separation) on glasscoverslips for measurement. In some cases, cells were grown in lowdensity overnight on glass coverslips. Potential blockers were testedfor ability to block current in the continued presence of AITC.

The IC₅₀ values presented in Table 1 were obtained from patch clampexperiments.

INCORPORATION BY REFERENCE

All publications and patents mentioned herein are hereby incorporated byreference in their entirety as if each individual publication or patentwas specifically and individually indicated to be incorporated byreference.

EQUIVALENTS

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

1. A method of treating an injury to the skin or respiratory tractresulting from exposure to a chemical warfare agent, the methodcomprising administering to a subject an effective amount of a compoundof Formula (I) or a salt thereof:

wherein R¹ is H; R² is H or C₁-C₆ alkyl; L is C(O)NR⁶; R³ isnitrogen-containing heteroaryl substituted with R⁷; R⁶ is H; R⁷ isheteroaryl substituted with 1-3 R⁸; each R⁸ is independently cyclyl orheterocyclyl; wherein said cyclyl or heterocyclyl can be substituted atone or more positions with halogen, alkyl, aralkyl, alkenyl, alkynyl,cycloalkyl, hydroxyl, amino, nitro, sulfhydryl, imino, amido, phosphate,phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether, alkylthio,sulfonyl, ketone, aldehyde, ester, a heterocyclyl, an aromatic orheteroaromatic moiety, —CF₃, or —CN; R⁹ is H; R¹¹ is H; R¹² is C₁-C₆alkyl; m is 1; and n is
 0. 2. A method of treating an injury to the skinor respiratory tract resulting from exposure to a chemical warfareagent, the method comprising administering to a subject an effectiveamount of a compound of Formula (I) or a salt thereof:

wherein R¹ is H or C₁-C₆ alkyl; R² is H; L is C(O)NR⁶; R³ isnitrogen-containing heteroaryl substituted with R⁷; R⁶ is H; R⁷ isheteroaryl substituted with 1-3 R⁸; each R⁸ is independently cyclyl orheterocyclyl; wherein said cyclyl or heterocyclyl can be substituted atone or more positions with halogen, alkyl, aralkyl, alkenyl, alkynyl,cycloalkyl, hydroxyl, amino, nitro, sulfhydryl, imino, amido, phosphate,phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether, alkylthio,sulfonyl, ketone, aldehyde, ester, a heterocyclyl, an aromatic orheteroaromatic moiety, —CF₃, or —CN; R⁹ is H; R¹¹ is H; R¹² is C₁-C₆alkyl; m is 1; and n is
 0. 3. The method of claim 1, wherein thecompound is administered orally, via intramuscular injection, ortopically.
 4. The method of claim 1, wherein the chemical warfare agentis tear gas.
 5. The method of claim 1, wherein the chemical warfareagent is chlorine.
 6. The method of claim 1, wherein the chemicalwarfare agent is mustard gas.
 7. The method of claim 1, wherein thesubject is a human.
 8. The method of claim 2, wherein the compound isadministered orally, via intramuscular injection, or topically.
 9. Themethod of claim 2, wherein the chemical warfare agent is tear gas. 10.The method of claim 2, wherein the chemical warfare agent is chlorine.11. The method of claim 2, wherein the chemical warfare agent is mustardgas.
 12. The method of claim 2, wherein the subject is a human.